A hepatocyte membrane-based culture system recapitulating the physiological microenvironment of hepatic stellate cells

Liver fibrosis is a progressive disease primarily driven by the activation of hepatic stellate cells (HSCs). Understanding the mechanisms regulating HSC activation requires an in vitro model that accurately recapitulates the hepatic microenvironment. In vivo, quiescent HSCs form direct contact with hepatocytes (Heps) through fine dendritic processes known as spines, which are essential for maintaining liver architecture and homeostasis. However, conventional extracellular matrix-based culture systems fail to reproduce these physiological cell-cell interactions. In this study, we aimed to establish a novel HSC culture platform using Heps plasma membrane (Hep-PM) as a substrate to mimic direct Heps-HSCs adhesion. Primary mouse and human HSCs cultured on Hep-PM retained dendritic, star-like morphologies characteristic of quiescent cells and exhibited markedly reduced expression of alpha-smooth muscle actin (α-SMA) and collagen type I alpha 1, key markers of HSC activation. Remarkably, Hep-PM also promoted the deactivation of activated HSCs, suggesting that both activation and reversion processes can be recapitulated in vitro. Furthermore, HSCs maintained on Hep-PM remained responsive to transforming growth factor β (TGF-β), indicating that quiescence was preserved without loss of activation potential. This system recreates the hepatic microenvironment, enabling dynamic and quantitative evaluation of HSC phenotypes. The Hep-PM platform offers a powerful tool for elucidating HSC regulatory mechanisms and screening antifibrotic compounds and may ultimately inform the design of Heps-mimetic therapeutics for liver fibrosis.<b>NEW & NOTEWORTHY</b> We developed a hepatocyte plasma membrane-based culture system that mimics in vivo cell-cell adhesion to regulate hepatic stellate cell (HSC) phenotypes. This platform maintains HSC quiescence by reducing activation markers and preserving a quiescent morphology, while allowing the observation of HSC activation in response to TGF-β treatment. It provides a physiologically relevant tool for studying HSC regulation and screening antifibrotic therapies, advancing in vitro modeling of liver fibrosis.